In the United States, 1.4 million patients per year undergo operations requiring arterial prostheses (Langer et al, Science 260:920-926 is (1993)). Approximately 100,000 patients per year require vascular bypass of small caliber arteries, but have no usable autologous artery or vein for grafting (Niklason, Science 286:1493-1494 (1999)). Hence, there is a pressing need for autologous vessels to treat atherosclerotic disease (Lefkowitz et al, Journal of American Medical Association 285:581-587 (2001)). Arteries have been successfully engineered from neonatal human smooth muscle cells (SMCs) (L'Heureux et al, FASEB J. 12:47-56 (1998)), and from porcine and bovine SMCs (Niklason et al, Science 284:489-493 (1999), Niklason et al, Journal of Vascular Surgery 33:628-638 (2001)). However, these approaches have not yet been translated to the growth of human arteries suitable for clinical use.
The inability of non-neonatal SMCs to form arteries in vitro may be due to their finite lifespan in culture. Specifically, arterial culture in vitro requires 45 to 60 population doublings (PD) of SMCs to produce a mechanically robust artery, while endothelial monolayer formation requires far fewer cell doublings (L'Heureux et al, FASEB J. 12:47-56 (1998), Niklason et al, Science 284:489-493 (1999), Niklason et al, Journal of Vascular Surgery 33:628-638 (2000)). However, non-neonatal human SMCs proliferate in vitro for only 15 to 30 PD before terminally growth arresting in a state termed senescence (Bierman, In Vitro 14:951-955 (1978), Bonin et al, Arteriosclerosis, Thrombosis, and Vascular Biology 19:575-587 (1999)). Hence, the limited lifespan of human SMCs represents a fundamental hurdle to the culture of autologous blood vessels.
Recently, numerous genetic approaches have been developed to extend the lifespan of human somatic cells. For instance, in several normal human somatic cells, expression of the hTERT gene (Nakamura et al, Cell 92:587-590 (1998)) (hTERT being the catalytic protein subunit of human telomerase reverse transcriptase) has been shown to reactivate telomerase, the enzyme that elongates chromosome-capping telomeres (Sedivy, Proceedings of the National Academy of Sciences USA 95:9078-9081 (1998)). Cellular lifespan was extended without induction of a cancerous phenotype (Jiang et al, Nature Genetics 21:111-114 (1999), Morales et al, Nature Genetics 21:115-118 (1999)), Yang et al, Journal of Biological Chemistry 274:26141-26148 (1999)).
The present invention results from studies demonstrating that expression of lifespan-extending genes in non-neonatal human cells (i.e., SMCs) extends the proliferative capacity of these cells. Approaches utilizing lifespan extension make possible the culture of robust arteries from human cells in vitro.